酶的分子设计、改造与工程应用

酶工程的研究已经发展到分子水平 ,在体外通过基因工程、化学、物理等手段改造酶分子结构与功能 ,大幅提高了酶分子的进化效率和催化效率 ,生产有价值的非天然酶。对酶工程学若干“热点”和前沿课题的研究、应用进行了概述 ,分析了国际上酶工程研究及应用技术、手段、方法 ,包括体外分子进化、核酶和抗体酶的设计、酶分子的定向固定化技术、酶蛋白分子的化学修饰、融合酶、人工合成及模拟酶等技术 ,并展望了酶工程的技术进步和应用的新进展。     

分子酶工程的研究进展

随着基因工程和蛋白质工程的进展和应用,酶工程在分子水平上的研究与应用也得到了迅猛发展。本着重介绍了酶基因克隆与异源表达、酶分子的定向改造和进化、融合蛋白与融合酶、酶的人工模拟(抗体酶、分子印迹技术)和端粒酶,综述了分子酶工程的研究进展、趋势及其应用。     

酶分子体外定向进化的研究方法*

酶分子体外定向进化不仅可大幅度提高酶分子的进化效率,短期内在实验室完成自然状态下需要千百万年的进化过程,还可使酶分子按照人们期望的特定目标进化,因此对酶工程今后的发展非常重要。本对酶分子体外定向进化的研究方法进行了归纳总结。     

植物分子系统分类学的新技术—VNTR及DNA指纹图

随着分子生物学技术日新月异的发展,许多先进的实验方法与较成熟的分析技术已开始进入植物系统分类学领域,使得这门学科逐步往分子水平迈进,可以直接对遗传变异和进化的分子基础DNA进行分析,以研究并揭示植物系统分类和进化的分子规律。继RFLP(Restriction Fragment Length Polymorphism,限制性酶切片段长度多态性)技术被植物系统分类学采纳并应用以来,最近,分子生物学     

DNA分子进化的研究基础和策略

从研究方法和技术手段等方面讨论了分子进化研究上的一些重大问题．阐述了基因的基本结构及DNA分子的进化方式：净化选择、中性进化和分化选择。探讨了利用DNA分子进化数据构建系统树的原理,评价了NJ、UPGMAM、MP和ML4种算法在构建进化树时的合理性与不足,以及进化树可靠性检验的方法,并对未来分子进化生物学的发展加以展望。     

植物几丁质酶的基因工程与分子生物学研究进展

植物几丁质酶具有广泛的生理活性,尤其在植物的抗病性方面具有重要作用,因而植物几丁喷酶基因工程或为目前抗真菌基因工程研究的热点。本介绍了植物几丁质酶基因结构的研究进展,综述了植物几丁质酶基因工程和微生物产几丁质酶转入植物的基因工程研究成果,概述了植物几丁质酶分子比较和分子进化研究,并展望了植物几丁质酶基因工程的应用前景。     

鞘翅目昆虫线粒体基因组研究进展

鞘翅目（Coleoptera）是世界上最具多样性的类群,具有很高的生态和形态多样性,这些多样性吸引了很多进化生物学家和分类学家的关注。随着分子生物学的发展,分子生物学技术广泛应用于鞘翅目系统学的研究,但随着研究的深入,简单的分子片段已经不能满足研究的需求,需要发掘更新的分子标记。近年来,线粒体全基因组已经成为鞘翅目分子系统学研究中很重要的分子标记之一,并广泛地应用于鞘翅目昆虫各个阶元的研究中。本文就鞘翅目线粒体全基因组的概况、研究进展及存在问题进行了总结和讨论。目前,鞘翅目线粒体基因组的研究主要包括物种线粒体基因组组成与结构、分子系统学和分子进化等方面。线粒体基因组在解决系统发育和进化方面表现出了很多的优越性,然而也存在着一些缺点,如序列难获得、基因类型单一、各基因进化速率不同、应用较局限等。     

融合酶的设计和应用研究进展

酶的分子改造和重新设计是解决酶催化工业应用瓶颈的重要途径。基于融合蛋白设计的融合酶技术是分子酶工程的一个研究热点,已逐渐应用于多功能酶和酶靠近效应的构建与控制研究中,显示出重要的理论和应用研究价值。文中对近年来融合酶的分子设计策略和应用研究的进展进行了综述。首先介绍了融合酶的概念和特点,并对最近研究中出现的融合酶构建策略进行了归纳总结,重点阐述了不同种类连接肽对融合酶的影响及其可能机理。同时,对目前融合酶的应用研究进行了归纳和讨论。最后,结合本实验室的研究,指出了融合酶领域的关键问题并对其发展方向进行了探讨和展望。     

分子生药学：一门新兴的边缘学科

分子生药学为生药学与分子生物学有机融合形成的新兴边缘学科。本文分析了分子生药学产生的背景、意义,介绍了分子生药学的研究内容,包括：生药分子鉴定、药用动植物的系统进化、药用动植物种质资源的评估及保存、药用动植物濒危机制及保护、药用植物活性成分的生物合成及调控、药用动植物的道地性及分子机理。认为分子生药学具有以下学科特色：（1）研究领域广泛,学科综合性很强;（2）次生代谢产物积累的研究贯穿分子生药学;（3）研究对象丰富多样,模式植物构建困难较大。简要介绍了分子生药学的常用技术方法,主要包括：DNA分析技术（分子杂交、分子标记技术、基因芯片、基因22程技术）、蛋白质分析技术、生物转化技术。指出分子生药学学科的发展呈现以下趋势：（1）学科理论体系进一步完善;（2）应用实践进一步加强;（3）生药资源永续利用的需求及技术发展导致特色领域成为热点,主要表现为分子鉴定稳步发展,biocoding成为分子鉴定的重要方向;次生代谢产物相关的功能基因组研究异军突起;基因组学、蛋白组学、代谢组学研究结果的整合和分析成为新热点;核心种质构建形成新思路和新方法。     

谈系统发生树建立的分子标准

随着进化生物学以及生物信息学的发展 ,研究不同物种间进化关系的方法也有了新的进展。其中分子进化方法使得系统发生树的建立更加简便和精确。几种系统发生树的建树方法及相关分子标准目前比较流行。。     

Directed enzyme evolution

Laboratory evolutionists continue to generate better enzymes for industrial and research applications. Exciting developments include new biocatalysts for enantioselective carbon-carbon bond formation and fatty acid production in plants. Creative contributions to the repertoire of evolutionary methods will ensure further growth in applications and expand the scope and complexity of biological design problems that can be addressed. Researchers are also starting to elucidate mechanisms of enzyme adaptation and natural evolution by testing evolutionary scenarios in the laboratory.     

2018酶工程专刊序言

酶工程是酶学与工程科学融合的综合性科学技术,是现代生物技术的支柱之一。为促进国内酶工程研究的发展,本期"酶工程专刊"集中展现了我国酶工程专家学者在酶工程领域所取得的最新进展。     

工业生物催化技术

以蛋白质酶的工程应用为核心的工业生物催化技术,被认为是生物技术继生物医药和转基因植物之后的第三次浪潮。它的发展与应用将对人类的工业化学过程带来根本的变革。工业生物催化的兴起与以下的两个关键技术因素有密切的关系：(1)蛋白质定向进化技术的出现,(2)基因组学和蛋白质组学的发展。探讨了工业生物催化技术的现状和发展趋势,并对我国如何发展该领域的基础和应用研究提出一些见解。     

工业酶研究中的计算化学方法

文中介绍用于工业酶研究特别是用于指导酶工程的主要计算化学方法,包括分子力学力场和分子动力学模拟、量子力学以及量子力学/分子力学结合模型、连续介质静电模型以及分子对接等。文中从以下两个角度分别概要地介绍这些方法:一是方法本身的基本概念、原始计算结果、适用条件和优缺点等;二是通过计算得到有价值的信息,指导突变体和突变库设计。     

Directed Computational Evolution of Quorum-Quenching Lactonases from the Amidohydrolase Superfamily

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Enzyme engineering: Reshaping the biocatalytic functions

Enzyme engineering is a powerful tool to fine-tune the enzymes. It is a technique by which the stability, activity, and specificity of the enzymes can be altered. The characteristic properties of an enzyme can be amended by immobilization and protein engineering. Among them, protein engineering is the most promising, as in addition to amending the stability and activity, it is the only way to modulate the specificity and stereoselectivity of enzymes. The current review sheds light on protein engineering and the approaches applied for it on the basis of the degree of knowledge of structure and function of enzymes. Enzymes, which have been engineered are also discussed in detail and categorized on the basis of their respective applications. This will give a better insight into the revolutionary changes brought by protein engineering of enzymes in various industrial and environmental processes.     

酶制剂研制的国内外进展和发展策略

简述了酶工程的研究范围,国际上酶制剂工业发展的研究,特点。概述了近年来我国酶制剂工业发展中存在的问题,包括产品结构,技术含量,研发能力。提出我国酶制剂工业今后发展的一些建议：充分发挥资源优势,加强现代生物技术的应用,调整酶制剂产品的结构。     

Protein evolution: The deciphering of latent facets — correlation of synthesis profiles of ribosomally directed proteins and enzyme directed peptides

Key facets pertaining to the evolution of proteins have been probed, using as springboard, the relevant data bases constructed from (i) 60 ribosomally directed proteins, whose 3D structures are known and having 10,000 residues and (ii) from 73 enzyme directed peptides, comprising of 524 residues. The preference profiles, both in terms of the choice of neighbours and the placement of the peptide bonds, have been delineated with respect to each of the 20 coded amino acids. By and large, the preference profile from both the sets are similar, thus giving importance to the nature of the side chains of the coded amino acids. The predictive power of the preference profile has been tested with good results, thus demonstrating the evidence of common preference pathways for peptide formation during evolution. The ribosomally directed protein synthesis, controlled by the genome, proceeds by the addition of single residues at a time. On the other hand, the enzyme directed peptide synthesis largely operates in a more energy conscious block mode, where each constituent of a large enzyme ensemble is engaged in precisely assembling the modules and transfering them to the adjacent one, thus realizing a sequence specific peptide synthesis. Of significance is the fact that, in spite of such divergence in assembly, predictions for neighbour preferences in ribosomally directed protein synthesis work well when applied to enzyme directed peptide synthesis. The findings here are significant since they provide (i) a clear picture of directed peptide synthesis in the absence of direct genomic control, (ii) evidence for the preferred formation of peptide bonds using protein templates, (iii) they also provide evidence for the presence of protein like structures, with catalytic activity, prior to the freezing of the genetic code arising from dominance of the information system and (iv) a logical approach to the evolution of a hierarchical pattern. Presented at the National Symposium on Evolution of Lite.     

Effects of helix and fingertip mutations on the thermostability of xyn11A investigated by molecular dynamics simulations and enzyme activity assays

Local conformational changes and global unfolding pathways of wildtype xyn11A recombinant and its mutated structures were studied through a series of atomistic molecular dynamics (MD) simulations, along with enzyme activity assays at three incubation temperatures to investigate the effects of mutations at three different sites to the thermostability. The first mutation was to replace an unstable negatively charged residue at a surface beta turn near the active site (D32G) by a hydrophobic residue. The second mutation was to create a disulphide bond (S100C/N147C) establishing a strong connection between an alpha helix and a distal beta hairpin associated with the thermally sensitive Thumb loop, and the third mutation add an extra hydrogen bond (A155S) to the same alpha helix. From the MD simulations performed, MM/PBSA energy calculations of the unfolding energy were in a good agreement with the enzyme activities measured from the experiment, as all mutated structures demonstrated the improved thermostability, especially the S100C/N147C proved to be the most stable mutant both by the simulations and the experiment. Local conformational analysis at the catalytic sites and the xylan access region also suggested that mutated xyn11A structures could accommodate xylan binding. However, the analysis of global unfolding pathways showed that structural disruptions at the beta sheet regions near the N-terminal were still imminent. These findings could provide the insight on the molecular mechanisms underlying the enhanced thermostability due to mutagenesis and changes in the protein unfolding pathways for further protein engineering of the GH11 family xylanase enzymes.